The small GTPase Gsp1p binds to the repeat domain of the nucleoporin Nsp1p.
The small GTPase Gsp1p of Saccharomyces cerevisiae and its homologue Ran play essential roles in several nuclear processes, such as cell-cycle progression, nuclear organization and nucleocytoplasmic traffic of RNA and proteins. Gsp1p/Ran is an abundant nuclear protein that interacts with different cytoplasmic and nuclear factors. Several of the previously identified Ran-binding proteins located at the nuclear-pore complex carry a specific Ran-binding domain. So far, direct interactions between the GTPase and other proteins have been mostly characterized in higher eukaryotes. Here we report that the yeast protein Gsp1p can directly bind to the nucleoporin Nsp1p in vitro. Nsp1p does not contain a Ran-binding domain and therefore represents a distinct type of nucleoporin that associates with Gsp1p. We demonstrate that the middle domain of Nsp1p is sufficient to mediate this interaction. Importantly, we show that a conserved cluster of positively charged amino acid residues of Gsp1p located at positions 142-144 is essential for the binding reaction. Thus we have identified Nsp1p as a new candidate protein located at the nuclear pore complex of the yeast S. cerevisiae that interacts directly with Gsp1p. We further demonstrate that both Gsp1p and Nsp1p are components of larger protein complexes in vivo, supporting the idea that the association between both proteins takes place in growing cells.
Stochaj
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U
Physiology Department, McGill University, 3655 Drummond Street, Montreal, PQ, Canada H3G 1Y6.
Héjazi
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M
Belhumeur
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P
D001665:Binding Sites; D002135:Calcium-Binding Proteins; D002467:Cell Nucleus / Q000378:metabolism; D003432:Cross-Linking Reagents; D005656:Fungal Proteins / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006150:Guanine Nucleotides / Q000378:metabolism; D046911:Macromolecular Substances; D020559:Monomeric GTP-Binding Proteins; D028861:Nuclear Pore Complex Proteins; D009687:Nuclear Proteins / Q000378:metabolism*; D017354:Point Mutation; D011233:Precipitin Tests; D011485:Protein Binding; D012091:Repetitive Sequences, Nucleic Acid; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins
D002135:Calcium-Binding Proteins; D003432:Cross-Linking Reagents; D005656:Fungal Proteins; C080324:GSP1 protein, S cerevisiae; D006150:Guanine Nucleotides; D046911:Macromolecular Substances; C062748:NSP1 protein, S cerevisiae; D028861:Nuclear Pore Complex Proteins; D009687:Nuclear Proteins; D029701:Saccharomyces cerevisiae Proteins; D019204:GTP-Binding Proteins; D020559:Monomeric GTP-Binding Proteins